Turpin et al. BMC Medical Genetics(2020) 21:204
https://doi.org/10.1186/s12881-020-01115-w

RESEARCH ARTICLE

Open Access

Associations of NOD2 polymorphisms with
Erysipelotrichaceae in stool of in healthy
first degree relatives of Crohn’s diseasesubjects
Williams Turpin1,2, Larbi Bedrani1, Osvaldo Espin-Garcia3,4, Wei Xu4, Mark S. Silverberg2, Michelle I. Smith1,
Juan Antonio Raygoza Garay1,2, Sun-Ho Lee1,2, David S. Guttman5,6, Anne Griffiths7, Paul Moayyedi8,
Remo Panaccione9, Hien Huynh10, Hillary A. Steinhart1,2, Guy Aumais11, Levinus A. Dieleman12, Dan Turner13, CCC
IBD GEM Project research team, Andrew D. Paterson4,14,15 and Kenneth Croitoru1,2*

Abstract
Background: Genetic analyses have identified many variants associated with the risk of inflammatory bowel disease(IBD) development. Among these variants, the ones located within the NOD2 gene have the highest odds ratio of
all IBD genetic risk variants. Also, patients with Crohn’s disease (CD) have been shown to have an altered gut
microbiome, which might be a reflection of inflammation itself or an effect of other parameters that contribute to
the risk of the disease. Since NOD2 is an intracellular pattern recognition receptor that senses bacterial
peptidoglycan in the cytosol and stimulates the host immune response (Al Nabhani et al., PLoS Pathog 13:
e1006177, 2017), it is hypothesized that NOD2 variants represent perfect candidates for influencing hostmicrobiome interactions. We hypothesized that NOD2 risk variants affect the microbiome composition of healthy
first degree relative (FDR) of CD patients and thus potentially contribute to an altered microbiome state before
disease onset.
Methods: Based on this, we studied a large cohort of 1546 healthy FDR of CD patients and performed a focused
analysis of the association of three major CD SNPs in the coding region of the NOD2 gene, which are known toconfer a 15–40-fold increased risk of developing CD in homozygous or compound heterozygous individuals.
Results: Our results show that carriers of the C allele at rs2066845 was significantly associated with an increase inrelative abundance in the fecal bacterial family Erysipelotrichaceae.
(Continued on next page)* Correspondence: ken.croitoru@sinaihealth.ca
1
Department of Medicine, University of Toronto, Toronto, ON, Canada
2
Zane Cohen Centre for Digestive Diseases, Mount Sinai Hospital, 600
University Avenue Room 437, Toronto, Ontario M5G 1X5, Canada
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Turpin et al. BMC Medical Genetics(2020) 21:204

Page 2 of 8

(Continued from previous page)

Conclusions: This result suggests that NOD2 polymorphisms contribute to fecal microbiome composition inasymptomatic individuals. Whether this modulation of the microbiome influences the future development of CDremains to be assessed.
Keywords: Fecal microbiota, Healthy human, Microbiome, rs2066844, rs2066845, rs2066847, NOD2, Inflammatory
bowel disease

Background
Crohn’s disease (CD) is a complex disease with a significant public health impact [1]. Several factors are known
to contribute to the risk of developing CD, but the exact
cause(s) remains to be identified. It is generally thoughtthat genetic and environmental factors play a crucial rolein the development of CD. Also, alterations in the gut
microbiome are described in patients with establishedCD [2]. NOD2 was the first susceptibility gene discovered for CD [3]. Since this discovery, genome-wide association studies (GWAs) have allowed the identificationof many other genetic variants associated with CD [4],including several additional variants located in the
NOD2 locus [4]. More precisely, three common variants
located in the NOD2 leucine-rich repeat domain havebeen identified in the European population. These are
R702W, G908R resulting from two cis-mutations in
rs2066844, rs2066845 and L1007fs, a frameshift mutation identified by rs2066847 insC, respectively [5, 6].
Individuals who are homozygous or compound heterozygous for these mutations in NOD2 have a 15–40-foldincreased risk of developing CD [5, 6].
Previous studies attempted to determine if NOD2
genotype is associated with any alteration in the composition of the intestinal microbiome in CD case-control
studies. However, those studies fail to distinguish between the possibilities that microbial changes are the result of inflammation present in these patients with CDor drug therapies [7]. Several studies have performed
such analysis using a GWAs approach in healthy individuals, but all failed to identify robust association looking
at NOD2 genotype association with microbial composition [8–10]. The conservative threshold for association
significance used in GWAs studies (p-value < 5 × 10− 8),may explain the absence of association. Alternatively,
healthy individuals from the general population may
have a relatively lower abundance of NOD2 variants associated with CD. For these two reasons, we addressedin a targeted fashion whether NOD2 CD risk alleles areassociated with microbial composition in healthy firstdegree relatives (FDR) of CD patients known to be genetically at higher risk to develop CD as compared togeneral population [11, 12]. We thus evaluated in a targeted fashion the association between three NOD2 CD
risk variants known to have the largest effect size for CD(rs2066844, rs2066845 and rs2066847) with the fecal microbial composition in a large cohort of 1546 healthy
FDRs of CD patients.

Methods
Patient recruitment

A total of 1546 subjects were recruited as part of the
Crohn’s and Colitis Canada Genetic Environmental Microbial (CCC-GEM) project between the years 2008 and2017. The study was approved by the Mount Sinai Hospital Research Ethics Board (Toronto - Managing center)and each participating recruitment center. All the participants or their guardians provided written consent before
their enrollment. Participants provided blood and a stool
sample for genetic and microbiome analysis, respectively.
Genotyping

DNA extraction from peripheral blood mononuclear
cells (PBMC) was performed using the Gentra Puregene
Blood Kit (Qiagen, CA, USA). Genomic DNA was quantified using Nanodrop®, and then dilutions were preparedat concentrations of 20 ng/μl and aliquoted into 96 wellreaction plates. SNPs genotyping was performed using
Illumina HumanCoreExome chip and ImmunoChip(Illumina, Inc. San Diego, CA) following the provider’srecommendations as previously described [8].
Genome imputation

Genotype imputation was performed on a merged
HumanCoreEXOME chip and ImmunoChip hard genotype data. We performed two imputations, the first one
based on the Haplotype Research Consortium (HRC)
panel, and the second based on the 1000 Genomes imputation panel of October 2014. The former was performed by directly uploading the hard genotype to the
HRC website (http://www.haplotype-reference-consortium.org/), while for the latter, IMPUTE2 (v. 2.3.0) [13,14] was used. Markers were imputed in 1 M bp intervals
with 750 K bp flanking regions with pre-phased haplotypes. Imputation results were quality controlled forminor allele frequency (MAF > 1%), missingness (< 5%),
imputation certainty (> 0.4), and imputation information(> 0.9). The three NOD2 SNPs were identified based on
their coordinates in the human genome build 37. The
two SNPs rs2066844, rs2066845 were captured from theTurpin et al. BMC Medical Genetics(2020) 21:204

HRC imputation, whereas the rs2066847 was present asa hard-genotyped SNP in HumanCoreEXOME chip.
PLINK v1.90b3.38 was used to extract the SNPs and to
calculate their MAF [15].
Taxonomic profiling of the gut microbiota

Stool samples were collected in FB Commode Specimen
Collector (Fisher Scientific, Waltham, MA) and put into
polypropylene vials (Starplex Scientific Inc., Etobicoke,ON) and frozen prior their shipping to the local study
sites where they were stored at − 80 °C. DNA extraction
from fecal bacteria was performed using the QIAamp
DNA Stool Mini Kit (Qiagen, Hilden, Germany). We
added a step to the manufacturer’s recommendations,which consists of physical disruption of the bacterial cellwall using ceramic beads. The V4 hypervariable regionof bacterial 16S ribosomal RNA (16S rRNA) was sequenced in paired-end mode (2 × 150 base pair) using
MiSeq platform (Illumina Inc., San Diego, CA, USA).
Paired reads were assembled using fastq-join software(ea-utils suite v1.1.2.537), resulting in 250 base pairs
amplicons. The latter were then processed using thequantitative insights into microbial ecology (QIIME
v1.9.0) pipeline using the default parameters. Operational taxonomic units (OTUs) were clustered at 97%
sequence similarity against Greengenes database v.13_8
using a closed reference approach. Samples with lessthan 10,000 reads after quality filtering were removed
from the analysis. Alpha diversity was expressed using
the Shannon index after rarefaction at a depth of 10,000
reads per sample. 16S sequences are available by thestudy accession number PRJEB14839 [8].
Statistical considerations

To assess the association between the NOD2 polymorphisms and the fecal microbiome, we used two approaches. In the first approach, we evaluated theindividual association of each of the three NOD2 risk
SNPs with the microbiome. In the second approach, we
stratified the cohort based on the presence of at least
one NOD2 risk variant. Hence we had a new variablewith two categories: subjects with at least one mutation(rs2066844, and/or rs2066845, and/or rs2066847) andsubjects with no mutations.
In the two approaches, we tested first the associationof the variables with the global composition of the
microbiome at the genus level using permutational
multivariate analysis of variance (PERMANOVA) on an
OTU table rarefied to 10,000 reads and using a BrayCurtis distance matrix. Then, the variables (NOD2 polymorphism and presence of at least one variant) were
tested for their association with every member of the different taxonomy levels from phylum to genus using a
two-part log-normal model as previously described [8,Page 3 of 816]. The model was applied to a non-rarefied OTU table
and corrected for age, sex and the total number of reads
per sample. All statistical analyses were performed using
R (v3.3.3). PERMANOVA was performed using theVegan package (V2.4–5), whereas the geeglm function of
the geepack package (v1.2.0–2) was used to carry out
the two-part log-normal model on each bacterial taxa.
The correction of multiple testing was performed by
calculating the effective number of uncorrelated taxa
using the approach described by Li and Ji [17]. Our taxonomy table contained 263 taxa from phylum to genus
level. The application of the Li and Ji [17] method
allowed us to identify 132 independent variables, bringing the raw p-value significance threshold to 3.8 × 10− 4.

Results
Frequency of NOD2 risk variants in healthy FDR of CDpatients

A cohort of 1546 asymptomatic FDRs of CD patients recruited between 2008 and 2017 as part of the Crohn’sand Colitis Canada Genetic Environmental Microbial(CCC-GEM) project. All study subjects were of Caucasian descent with a mean age ± standard deviation of
19.4 ± 7.9 [6-35] were included in this study including
848 Female (See methods section). We calculated the
MAF of the rs2066844 (R702W), rs2066845 (G908R)
and rs2066847 (l1007fs) in our population of healthy
FDRs of CD patients and compared this to those recorded in the 1000 genome project (self-reported healthy
individuals) (see Table 1). The MAF of each of the three
NOD2 risk SNPs were numerically higher in our cohort
than reported in the 1000 genome project general
healthy population. This increase in SNP frequency suggests that our cohort is enriched in NOD2 risk SNPs.
Association of the individual NOD2 risk SNPs with themicrobial composition and diversity of the microbiome in
FDRs

To understand the associations between the three individual NOD2 variants and the significance in clustering
patterns of the microbiome composition, we performed
a PERMANOVA analysis; this showed no significant association (Table 2). Similarly, the group with at least one
out of the three NOD2 variants compared with individuals without any NOD2 variant showed no significant
difference in clustering patterns of stool microbiome
composition. The alpha diversity, as represented by the
Shannon index, also showed no significant association
with each of the individual NOD2 variants, rs2066844,
rs2066845 or rs2066847, respectively. The group with atleast one mutation in the three NOD2 risk SNPs compared to the group with no variants also showed no significant difference in alpha diversity.

Turpin et al. BMC Medical Genetics(2020) 21:204

Table 1 Minor allele frequency of NOD2 risk SNPs in healthy
FDR of CD patients in comparison to the MAF in the 1000
genome project
SNPs

MAF in healthy
FDR of CD

MAF in the 1000
genomes projectrs2066844

8.9%

1%

rs2066845

3.1%

< 1%

rs2066847

4.2%

< 1%

MAF: Minor allele frequency, SNPs: single nucleotide polymorphisms. MAF
from the 1000 genomes project was obtained using Ensembl database

Association between NOD2 polymorphisms and specificfecal bacterial taxa and diversity

We assessed the association of each of the three NOD2 variants with each fecal bacterial taxon from phylum to genus
level of the taxonomy. The rs2066845 was significantly associated with the family Erysipelotrichaceae (estimate ±standard deviation; 0.39 ± 0.10, p-value = 7.14 × 10− 5)(Fig. 1). The abundance of this taxon was significantly increased in the groups with at least one mutation in
rs2066845 (the heterozygote and the minor homozygote)compared to groups with the null mutation. However, noneof the genera of the Erysipelotrichaceae family appeared tosignificantly contribute to the effect observed at the familytaxonomic level (Supplementary Table 1). No significant associations were observed for rs2066844 or rs2066847 withmicrobiome taxa (Supplementary Tables 2 and 3, respectively). When assessing the effect of binning the three
NOD2 variants, no significant associations were observed
between the presence of at least one mutation within thethree NOD2 variants with any of the individual taxa
(Supplementary Table 4).

Discussion
The NOD2 gene was the first found to be directly associated with CD risk through three main mutations occurring in the leucine-rich repeat region of the gene. Since
this gene is a pattern recognition receptor and is involved in host-microbe interactions, a few studies haveshown that NOD2 polymorphisms are involved in shaping the microbiome in IBD patients [18–21]. However,most of these studies were case-control studies, and as
Table 2 Association between the three NOD2 polymorphisms
and the clustering patterns of the fecal microbiome
composition
Variants

PERMANOVA p value

rs2066844

0.44

rs2066845

0.07

rs2066847

0.10

At least one NOD2 mutationa

0.75

a

in rs2066844 and/or rs2066845 and/or rs2066847

Page 4 of 8such, the inflammation process can affect microbiome
composition regardless of the NOD2 genotype [19–21].
Our study cohort of healthy FDRs of CD patients permits to minimize some of these potential confounding
factors as they are asymptomatic, do not take any antiinflammatory or anti-bacterial medication, while having
a high genetic risk of developing CD and carrying ahigher frequency of CD associated mutations. As such,
we assessed the association of three major NOD2 mutations with the composition of the gut microbiome in thiscohort. We found that this cohort was enriched in
NOD2 variants as reflected by higher MAFs of the CDrisk SNPs, rs2066844 (R702W), rs2066845 (G908R), andrs2066847 (l1007fs) as compared to the general population (Table 2) confirming the findings of our previous
study of healthy first-degree relatives [22]. Hence thiscohort provides a higher prevalence of the NOD2 polymorphisms and thus is more powered than cohorts ofhealthy individuals from the general population in order
to assess the potential association of NOD2 SNPs withgut microbiome composition in healthy individuals.
Our results showed that one of the NOD2 risk allele(rs2066845) is associated with the increased relative
abundance of Erysipelotrichaceae family. Little is known
regarding Erysipelotrichaceae family in terms of diseaseassociations [23]. There is data to suggest that Erysipelotrichaceae is associated with metabolic disorders inhumans [23]. Another study showed that species within
Erysipelotrichaceae family are positively linked to inflammation in patients who had chronic HIV infection [19].
This taxon was decreased in early CD onset [20], andpatients who experienced recurrence of CD had significantly lower levels of Erysipelotrichaceae [21]. Studies
have shown inconsistent alterations in Erysipelotrichaceae family relative abundance in patients with established CD [7, 20, 21, 23]. In a recent report,
Erysipelotrichaceae family was found to be increased inpatients with CD patients with ileal disease compared tothose with ileocolonic disease [24], which may be consistent with the fact that NOD2 is strongly associatedwith ileal CD [25]. Also, this taxon was found to behighly coated by IgA relative to other members of thegut microbiota in IBD patients [26]. It includes strictly
anaerobic or aerotolerant organisms, and members of
the Erysipelotrichaceae family can produce palmitic, stearic and oleic acid, fatty acids predominantly [27]. Overaccumulation of tissue palmitic acid results in dyslipidemia, hyperglycemia, increased ectopic fat accumulation,
increased inflammatory tone via toll-like receptor 4 [28]as well as pro-inflammatory adipokines mRNA expression in macrophages [29]. Also, stearic acid was shownto stimulate pro-inflammatory cytokine production [30].
It remained to be shown how the changes in this taxon
contribute to the risk of CD in this population.

Turpin et al. BMC Medical Genetics(2020) 21:204

Page 5 of 8

Fig. 1 Erysipelotrichaceae family is associated with rs2066845 genotype in 1546 healthy subjects. The x-axis corresponds to the rs2066845
genotype in the cohort. The y-axis corresponds to the log-transformed relative abundance of Erysipelotrichaceae. Each dot represents the value
for a given individual. The lines represent the first, second, and third quartiles. Carrier of the C allele are enriched in the relative abundance of
Erysipelotrichaceae (p-value = 7.14 × 10− 5)

The other two NOD2 risk variants (rs2066844 andrs2066847) showed no significant associations with the
fecal bacterial composition. A previous study detected
an association of NOD2 risk allele dosages (comprised of
combined rs104895431, rs104895467, rs2066844,
rs2066845, rs5743277, rs5743293) with an increase ofEnterobacteriaceae from biopsy sample in a cohort of474 IBD affected patients [31]. However, the alteredmicrobiome described in IBD patients is characterized
by an increase in Proteobacteria species such as Escherichia coli (a species belonging to Enterobacteriaceae family) [2, 32]. Thus the increase of Enterobacteriaceae
previously observed might be related to inflammation itself and not necessarily NOD2 genotype. Other studiesof large cohorts of healthy individuals have failed toshow any significant association of NOD2 genotype withmicrobiome composition [7–10, 33]. In these GWAS,
only one study comprised of 1514 individuals showedthat the NOD2 locus was associated with the enterobactin biosynthesis pathway, which is highly correlated with
Escherichia coli abundance [10]. Our analysis showedthat all the three tested NOD2 variants had a consistent
direction of the association for which NOD2 variants

had an increased Enterobacteriaceae despite not being
significant. Thus the association of Enterobacteriaceae
and NOD2 locus polymorphisms remains unclear.
The association between host genetics with microbial
composition might be better detected in biopsy samples
compared to fecal samples. In fact, in a study comparing the tissue associated bacteria in the ileum to the
fecal bacteria in CD patients carrying the rs2066847
polymorphism or not showed that significant differences in microbial composition were only observed in
the biopsy microbiome [34]. Recent GWAs studies of
IBD subjects report that NOD2 variants are strongly associated in particular with ileal CD compared to colonic CD [25]. Thus, this may, in part, explain theweaker signal observed in fecal samples compared tobiopsy samples in the ileum [35, 36].

Conclusion
In conclusion, this study is one of the largest studiesof healthy subjects examining the specific associationof the NOD2 genotype with the healthy human gut
microbiota. We found that the NOD2 genotype has
no significant influence on the overall composition ofTurpin et al. BMC Medical Genetics(2020) 21:204

stool microbiota, suggesting that NOD2 polymorphism
is not a major determinant of the overall compositionof the microbiome in this healthy at-risk population.
However, in our targeted analysis of the microbiota,
we found that rs2066845 is significantly associatedwith an increased relative abundance of the Erysipelotrichaceae family. It remains possible that the effectof NOD2 polymorphisms on microbiome composition
are limited under stable homeostatic conditions, andmight be more enhanced after the disease onset, i.e.
in the presence of active inflammation [31, 34, 37], or
may be related to a specific location such as ileal CD.
It remains possible that increased risk of developing
CD contributed by NOD2 genotype is in part related
to alteration of particular taxa such as Erysipelotrichaceae family in healthy individuals.

Supplementary information
Supplementary information accompanies this paper at https://doi.org/10.
1186/s12881-020-01115-w.
Additional file 1 Supplementary Table 1. Association of rs2066845
with bacterial taxa in the GEM cohort. Supplementary Table 2.
Association of rs2066844 with bacterial taxa in the GEM cohort.
Supplementary Table 3. Association of rs2066847 with bacterial taxa in
the GEM cohort. Supplementary Table 4. Cumulative NOD2 variants
association with bacterial taxa in the GEM cohort.
Abbreviations
CD: Crohn’s disease; IBD: Inflammatory bowel disease; FDR: Healthy first
degree relative; GWAs: Genome-wide association studies; CCC-GEM: Crohn’sand Colitis Canada Genetic Environmental Microbial; PBMC: Peripheral bloodmononuclear cells; PERMANOVA: Permutational multivariate analysis ofvariance; HRC: Haplotype Research Consortium; OTUs: Operational taxonomic
units; MAF: Minor allele frequency; SNP: Single nucleotide polymorphisms;
NOD2: Nucleotide-binding oligomerization domain-containing protein 2; 16S
rRNA: 16S ribosomal RNA; QIIME: Quantitative insights into microbial ecology
Acknowledgments
We thank the members of the CCC GEM Global Project Office. The CCC IBD
GEM Project research team is composed of: Maria Abreu, Paul Beck, Charles
Bernstein, Kenneth Croitoru, Leo Dieleman, Brian Feagan, Anne Griffiths,
David Guttman, Kevan Jacobson, Gilaad Kaplan, Denis O. Krause*(*deceased), Karen Madsen, John Marshall, Paul Moayyedi, Mark Ropeleski,
Ernest Seidman*(* deceased), Mark Silverberg, Scott Snapper, Andy Stadnyk,
Hillary Steinhart, Michael Surette, Dan Turner, Thomas Walters, Bruce
Vallance, Guy Aumais, Alain Bitton, Maria Cino, Jeff Critch, Lee Denson,
Colette Deslandres, Wael El-Matary, Hans Herfarth, Peter Higgins, Hien Huynh,
Jeff Hyams, David Mack, Jerry McGrath, Anthony Otley, and Remo Panancionne. The CCC GEM Project recruitment site directors include Maria Abreu,
Guy Aumais, Robert Baldassano, Charles Bernstein, Maria Cino, Lee Denson,
Colette Deslandres, Wael El-Matary, Anne M. Griffiths, Charlotte Hedin, Hans
Herfarth, Peter Higgins, Seamus Hussey, Hien Hyams, Kevan Jacobson, David
Keljo, David Kevans, Charlie Lees, David Mack, John Marshall, Jerry McGrath,
Sanjay Murthy, Anthony Otley, Remo Panaccione, Nimisha Parekh, Sophie Plamondon, Graham Radford-Smith, Mark Ropeleski, Joel Rosh, David Rubin, Michael Schultz, Ernest Seidman, Corey Siegel, Scott Snapper, Hillary Steinhart,
and Dan Turner.
Authors’ contributions
Conceived and designed the analysis: WT, LB, WX, MIS, CCC IBD GEM Project
research team, KC, MSS, DSG, AG, PM, RP, HH, HAS, GA, LAD, DT, ADP.
Collected the data: MSS, DSG, AG, PM, RP, HH, HAS, GA, LAD, DT, CCC IBD
GEM Project research team, KC. Contributed data or analysis tools: WT, LB,

Page 6 of 8

OEG, WX, MIS, JARG, SHL, PM, ADP, KC. Performed the analysis: WT, LB, OEG.
Wrote the paper: WT, LB, OEG, JARG, SHL, KC. The authors read andapproved the final manuscript
Funding
This study was supported by grants from Crohn’s and Colitis Canada Grant#CCC-GEMIII, Canadian Institutes of Health Research (CIHR) Grant#CMF108031 and the Helmsley Charitable Trust. Williams Turpin is a former
recipient of a Postdoctoral Fellowship Research Award from the CIHR
Fellowship/ Canadian Association of Gastroenterology (CAG)/ Ferring
Pharmaceuticals Inc. and of a fellowship from the Department of Medicine,
Mount Sinai Hospital, Toronto. Williams Turpin is partially supported by the
Biocodex Microbiota Foundation. Sun-Ho Lee, Juan Antonio Raygoza Garay,
are recipient of fellowship from the Department of Medicine, Mount Sinai
Hospital, Toronto. Mark Silverberg is supported in part by the Gale and Graham Wright Chair in Digestive Diseases. Kenneth Croitoru is partially supported by a Canada Research Chair in Inflammatory Bowel Diseases. Osvaldo
Espin-Garcia is a recipient of a CIHR STAGE fellowship.
Availability of data and materials
16S sequences are available by the study accession number PRJEB14839(NCBI BioProject database); ENA-SUBMISSION: ERA675979 [9]. Requests for
raw and analyzed data should follow the instruction given at http://www.
gemproject.ca/data-access/. All submissions will be reviewed by the GEM
Project Operating Committee to ensure that the requested samples/data will
not interfere in any way with the intended GEM Project analysis of thenested cohort as per the original GEM Project Study Design and is not a duplication of analysis already ongoing. Those proposals meeting this evaluation will be distributed to all members of the GEM Project Steering
Committee (GPSC) for review and open discussion. This review will focus on
the global scientific merit of the proposal. This review will assess the basic
scientific merit and the availability of requested samples and data, ensuring
there is no compromise of the original intent of the GEM project. It would
be of value to contact a member of the Steering Committee who could help
sponsor your application. Those projects achieving majority vote of approvalat the GPSC will be informed that the GEM Project will provide a letter of
support stating that the requested samples or data will be made available tothe applicants once the applicant receives funding from a granting agency
that applies an independent peer review process to the proposal. The criteria
to be used for review of all submissions will include the “scientific relevance”
of the proposal and the judged availability of biological material requested.
The budget to be requested from a funding agency, must allow for any expenses in processing samples or in setting up the appropriate queries of thedatabase. The intent is to allow sufficient time for applicants to consider submission for funding opportunities.
Ethics approval and consent to participate
All subjects and/or their guardians gave written informed consent toparticipate in the study. The study was approved by the Mount Sinai
Hospital Research Ethics Board (Toronto – Managing center) and local
centers (07–0322-E). The local centers are comprised of: the Conjoint
Health Research Ethics Board (Calgary), The Research Ethics Office(Edmonton), the Nova Scotia Health Authority (Halifax), the Hamilton
Integrated Research Ethics Board (Hamilton), the Queen’s University
Health Sciences and Affiliated Teaching Hospitals Research Ethics Board(Kingston), Western’s Research Ethics Boards (London, ON), the ResearchCentre of the Sainte-Justine University Hospital Research Ethics Board(Ste-Justine), McGill Research Ethics Boards (McGill), the Office of Research Ethics and Integrity (Ottawa), the Interdisciplinary Committee on
Ethics in Human Research (St John’s), the Clinical Research Ethics andRegulatory Compliance Office (Sick Kids Hospital), the UBC Office of Research Ethics (Vancouver), the Biomedical Research Ethics Board (Winnipeg), the comité d’éthique de la recherche (Hôpital du St-Sacrement),
the University of Saskatchewan Research Ethics Board (Saskatoon), theSunnybrook’s research ethics board (Sunnybrook), the MUHC Centre forApplied Ethics (Hôpital Maisonneuve-Rosemont), the UBC Providence
Health Care Research Ethics Board (Rhode Island), the University Health
Network Research Ethics Board (Toronto General Hospital), Comité d’éthique de la recherche du CIUSSS de l’Estrie (Sherbrooke), the Institutional
Review Board (Connecticut), The DHMC Clinical Ethics Committee (Dartmouth-Hitchcock Medical Center), the Helsinki Committee (ShaareTurpin et al. BMC Medical Genetics(2020) 21:204

Zedek), the Helsinki Committee of Rambam (Rambam), the Tel Aviv
Sourasky Medical Center Institutional Review Board (Tel Aviv Sourasky
Medical Center), the Helsinki Committee of Soroka (Soroka Medical Center), the Helsinki Committee of Baruch Padeh Medical Center, Poria (Poria
Medical Center), Kaplan Medical Center Helsinki Committee, the Kaplan
Medical Center, Rehovot (Kaplan), the Helsinki Committee of Tel Hashomer (Tel Hashomer Hospital), the Helsinki Committee of Emek (Emek),
the Assaf Harofeh Medical Center Helsinki Committee, Assaf Harofeh
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Center), the Helsinki Committee (Beilinson\Schneider), the Helsinki Committee of Maccabi (Maccabi Hashalom), the Helsinki Committee of Wolfson (Wolfson), the BSD/UCMC Institutional Review Boards at the
University of Chicago (Chicago), the Miami Research Ethics and Integrity
Program (Miami), the Lady’s Children’s Hospital Crumlin Research Ethics
Committee (Ireland), the Hospital Research Ethics Committee (Ireland),the West of Scotland Research Ethics Committee 4 (Royal Hospital for
Sick Children - Edinburgh), the Convener of the University Research Ethics Committee (Ninewells - Dundee), the West of Scotland Research Ethics Committee 4 (Western General Hospital - Edinburgh), the West of
Scotland Research Ethics Committee 4 (Yorkhill - Glasgow), the West of
Scotland Research Ethics Committee 4 (The Glasgow Royal Infirmary), the
NRES Research Ethics Committee (Aberdeen Royal Infirmary the West of
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Hospital), the South Birmingham Local Research Ethics Committee(Queen Elizabeth Hospital Birmingham), the HRA National Research Ethics
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Coventry), the North West - Greater Manchester Central Research Ethics
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Foundation Trust (The Royal Devon and Exeter Hospital), the Southampton & South West Hampshire local research ethics committee (Southampton General Hospital), the Central University Research Ethics
Committee (The John Radcliffe Hospital), the National Research Ethics
Service (Addenbrooke’s Hospital), the NHS Research Ethics Committee(Norfolk and Norwich University Hospitals), the Clinical Ethics Committee
at Barts and the London NHS Trust (St Bartholomew’s Hospital), the Westof Scotland Research Ethics Committee 4 (University Hospital Crosshouse), the Great Ormond Street Clinical Ethics Service (Great OrmondStreet Hospital), t NHS Lothian (Alder Hey Children’s Hospital), the Westof Scotland Research Ethics Committee 4 (Forth Valley Royal Hospital),
the Human Research Protection Office (CHIP Pittsburgh), North West Greater Manchester East Research Ethics Committee (Salford royal hospital), the Memorial Health Services Research Council (MemorialCare’s
Long Beach Medical Center), the Bnai-Zion Medical Center Helsinki Committee (Bnai Zion Medecal Center), the Institutional Review Board (Children’s Hospital Los Angeles), the West of Scotland Research Ethics
Committee 4 (Gartnavel General Hospital), the Southern District Health
Board and the Dunedin School of Medicine (Dunedin Hospital), the Central University Research Ethics Committee (John Radcliffe Hospital), theCode of Ethics and Corporate Compliance Program of Atlantic Health
System (New Jersey), the University Ethics of Research Committee (University Hospitals Bristol), the UC Irvine Interdisciplinary Center for the Scientific Study of Ethics and Morality (University of California Irvine), the
NHS Research Ethics Committees (Royal Free Hospital), the Wirral University Teaching Hospital NHS (Arrowe Park Hospital), the Tallaght University
Hospital / St. James’s Hospital Joint Research Ethics Committee (StJames’s Hospital), the Auckland Health Research Ethics Committee (Auckland District Health Board), the Waikato DHB (Waikato District Health
Board), the Helsinki Committee (Emek Medical Center), the Institutional
Review Board (Children’s Hospital of Philadelphia), the Helsinki Committee (Karmel Medical Centre), the NHS Research Ethics Committees (Barnet and Chase Farm NHS Hospitals), the NHS Research Ethics
Committees (New Victoria Hospital), the QIMR Berghofer Medical Research Institute-Human Research Ethics Committee (QIMR Berghofer
Medical Research Institute), the Helsinki Committee (French Hospital

Page 7 of 8

Nazereth), the Regional Ethics Review Board in Stockholm (Örebro University Hospital), the Regional Ethics Review Board in Stockholm (Karolinska University Hospital), and Regional Ethics Review Board in
Stockholm (Linköping University Hospital).
Consent for publication
Not applicable.
Competing interests
All authors disclose no potential conflicts (financial, professional, or personal)
that are relevant to the manuscript.
Author details
Department of Medicine, University of Toronto, Toronto, ON, Canada. 2Zane
Cohen Centre for Digestive Diseases, Mount Sinai Hospital, 600 University
Avenue Room 437, Toronto, Ontario M5G 1X5, Canada.
3
Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Toronto,
Ontario, Canada. 4Division of Biostatistics, Dalla Lana School of Public Health,
University of Toronto, Toronto, Ontario, Canada. 5Department of Cell &Systems Biology, University of Toronto, Toronto, Ontario, Canada. 6Centre for
the Analysis of Genome Evolution & Function, University of Toronto, Toronto,
Ontario, Canada. 7Division of Gastroenterology, Hepatology and Nutrition,
Department of Paediatrics, The Hospital for Sick Children, Toronto, Ontario,
Canada. 8Department of Medicine, McMaster University, Hamilton, Ontario,
Canada. 9Inflammatory Bowel Disease Clinic, Division of Gastroenterology
and Hepatology of Gastroenterology, University of Calgary, Calgary, Alberta,
Canada. 10Department of Pediatrics, University of Alberta, Edmonton, Alberta,
Canada. 11Hôpital Maisonneuve-Rosemont, Department of Medicine,
Montreal University, Montreal, Quebec, Canada. 12Division of
Gastroenterology and CEGIIR, Department of Medicine, University of Alberta,
Edmonton, Alberta, Canada. 13Department of pediatric GI, Shaare Zedek
Medical Center, 91031 Jerusalem, Israel. 14Division of Epidemiology, Dalla
Lana School of Public Health, University of Toronto, Toronto, Ontario,
Canada. 15Genetics and Genome Biology, The Hospital for Sick Children
Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada.
1

Received: 6 January 2020 Accepted: 31 August 2020

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